The COUP-TFs are orphan nuclear receptors proven to play important roles in controlling diverse aspects of growth, development, differentiation and homeostasis in several organisms. The COUP-TFs are regulators of gene expression and belong to the steroid/thyroid hormone receptor superfamily of ligand-activated transcription factors. The pattern and timing of expression of an orphan nuclear receptor gene defines the sphere of direct influence of a putative ligand signaling pathway within its target tissues and cells. The spatial and temporal expression pattern of COUP-TFs during inner ear organogenesis suggest that they may play a critical role in regulating processes that lead to the formation of a functional inner ear. Indeed, COUP-TFI loss-of-function mutants are deaf, having defects in development of the cochlea and sensory maculae of the vestibule. This suggests that COUP-TFI is an essential gene for the development of both hearing and balance functions in the mouse. To better understand the role of COUP-TFI in hearing and balance this grant will focus on the following aims. 1. Characterize defects in COUP-TFI mutants with respect to a detailed expression analysis of COUP-TFI during development and differentiation of inner ear structure and the brainstem auditory and vestibular nuclei. 2. Determine the mechanism of COUP-TFI action using chimeric embryo analysis to ascertain whether COUP-TFI acts in a cell-autonomous or non-autonomous manner in the ear. 3. Identify COUP-TFI downstream targets during inner ear development by subtraction library screens of cochlea and vestibule cDNA libraries. The completion of these aims will provide substantial insight on this novel regulator of inner ear morphogenesis and will identify genes required in its signaling pathway for development of hearing and balance.